Method for detecting single-chain DNA base mutation by gold nanometer particle

A gold nanoparticle, single-stranded technology, which is applied in the field of detecting single-stranded DNA base mutations using gold nanoparticles, can solve the problems of complicated operation, time-consuming and laborious, high cost, etc., and achieves simplified detection steps, easy mass preparation, and reduced cost. The effect of testing costs

Inactive Publication Date: 2008-09-10
XIAMEN UNIV
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Problems solved by technology

Some of these methods have poor specificity and low sensitivity, some are complicated to operate, time-consuming and labor-intensive, and some are prone to false positive results and high-cost detection, so they cannot meet the needs of clinical analysis of a large number of samples
DNA sequencing is the most direct and accurate method, but it requires special instruments, is time-consuming, laborious, expensive, and inconvenient to use in general units

Method used

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  • Method for detecting single-chain DNA base mutation by gold nanometer particle
  • Method for detecting single-chain DNA base mutation by gold nanometer particle
  • Method for detecting single-chain DNA base mutation by gold nanometer particle

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Embodiment 1

[0023] The detection method was applied to detect the mutation hotspot at codon 273 of p53 gene, the base G was mutated to the base T (see references: Rangel-Lopez A, Maldonado-Rodriguez R, Salcedo-Vargas M, et al.Low density DNAmicroarray for detection of most frequent TP53 missense point mutations. BMC Biotechnol, 2005, 15(8): 1-13), the specific operation steps are as follows:

[0024] 1) Mix 94mL of ultrapure water, 1mL of 1% chloroauric acid solution and 5mL of 1% trisodium citrate solution at room temperature, immediately put it in a microwave oven, heat it on high heat to make it boil, then adjust to medium heat , continue to heat for 8min to maintain the boiling state, cool to room temperature naturally, then centrifuge, concentrate, add water to redisperse, prepare colloidal gold with a particle size of 15nm, the color is dark red and translucent, the prepared colloidal gold TEM electron microscope picture see figure 1 ,Depend on figure 1 It can be seen that the part...

Embodiment 2

[0031] The detection method was applied to detect the mutation hotspot at codon 248 of the p53 gene, and the base G was mutated to the base A (see references: Rangel-Lopez A, Maldonado-Rodriguez R, Salcedo-Vargas M, et al.Low density DNAmicroarray for detection of most frequent TP53 missense point mutations. BMC Biotechnol, 2005, 15(8): 1-13), the specific operation steps are as follows:

[0032]1) Mix 96 mL of ultrapure water, 1 mL of 1% chloroauric acid solution and 3 mL of 1% trisodium citrate solution at room temperature, immediately put it in a microwave oven, add high heat to boil, and then adjust to medium heat , continue to heat and keep boiling for 8min, cool to room temperature naturally, then centrifuge, concentrate, add water to redisperse, and prepare colloidal gold with a particle size of 25nm, bright red and translucent.

[0033] 2) Take 10 fmol of wild-type single-stranded DNA and add it to 50 μL of colloidal gold. After mixing, stand still at room temperature ...

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Abstract

A method for using gold nanoparticles to detect single-stranded DNA base mutation relates to a detection of gene mutation and is provided. Colloidal gold is prepared to react with the single-stranded DNA to form the colloidal gold and a single-stranded DNA composite system; hydrochloride is added and the color of the composite system turns from red to purple or blue. Whether the single-stranded DNA mutates or not is judged by making use of the sequence differences between a wild-type single-stranded DNA and a mutant single-stranded DNA and through the final color changes of the composite system or the differences of the positions of the maximum absorption peak red shift of ultraviolet-visible absorption spectra. Both the single-stranded DNA and the gold nanoparticles can detect single base mutation without any decoration or special device; the lowest detection limit is 10 fmol and detection results can be given within 30 minutes. The method has the advantages of convenience, rapidness, low cost, high sensitivity, high specificity, strong practicality and so on and has big clinical application potential.

Description

technical field [0001] The invention relates to the detection of gene mutation, in particular to a method for detecting base mutation of single-stranded DNA by using gold nanoparticles. Background technique [0002] Gene mutation refers to the phenomenon of mutation, deletion or insertion of bases at one or more sites in the genomic DNA sequence. Gene mutation detection plays a very important role in biomedical research and can be used as an early clinical detection index for hereditary diseases, tumors, etc., and is of great significance for the timely detection and treatment of diseases ([1] Ellegren H. microsatellite DNA sequences. Nature Genetics, 2000, 24(4):400-402). [0003] Traditional gene mutation detection methods generally rely on flat-plate gel electrophoresis to analyze DNA conformation or melting properties, and then judge whether there is a mutation based on the difference in mobility during electrophoresis. Some of these methods have poor specificity and l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/33G01N21/78C12Q1/68
Inventor 孙莉萍张建锋李辉张其清王秀燕张召武
Owner XIAMEN UNIV
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